Analysing the genetic burden of rare and potentially damaging variants in schizophrenia risk-locus xq28,distal using high-throughput, multiplex targeted sequencing

Sex-specific differences in schizophrenia (SCZ) are pervasive and have been reported for various aspects of the disease. Previous authors have therefore hypothesized that genetic risk factors on the X chromosome play a role in the complex interplay that leads to the development of this multifactorial and highly polygenic disorder. Nevertheless, few studies specifically targeted the investigation of these sex-specific genetic risk factors. In recent years, large-scale systematic genetic studies in SCZ have broadened the spectrum of genetic variation contributing to the disease and identified, among others, several X-chromosomal risk-associated loci. The world´s largest copy number variant (CNV) analysis for SCZ reported the first potential risk-associated CNV on the X chromosome. Duplications in Xq28,distal confer risk to SCZ in both sexes and span eight different protein-coding genes. Due to the shared genomic breakpoints of the duplications described to date, the issue of which gene/s at this locus is/are relevant in terms of SCZ pathogenesis remains unclear. In order to detect an enrichment of rare and potentially damaging variants in possible disease-relevant genes, the objective was to provide a detailed analysis of the genetic burden within the Xq28,distal locus using high-throughput, multiplex targeted sequencing. All coding regions of the implicated genes were sequenced in a large, European SCZ case-control cohort (n= 1935 patients with SCZ and n=1905 population-based controls) using the single molecule molecular inversion probe method on an Illumina HiSeq2500. To identify rare and potentially damaging variants, the analyses focused on non-synonymous single-nucleotide variants and Indels with a minor allele frequency ≤ 0.1% and a Combined Annotation Dependent Depletion score ≥ 20. The burden analysis was performed in a gene-wise manner and with sex as a covariate using the X-chromosomal model of the Optimal Unified Sequence Kernel Association Test adjusting for possible sex-specific effects. A total of n=73 independent variants passed the applied quality control criteria. After filtering for rare and potentially damaging variants, a total of n= 13 independent variants, as identified in four cases and eleven controls, were subjected to the consecutive burden analysis. For the genes within the Xq28,distal locus, no significant single-gene association was identified (smallest value in BRCC3 with p=0.103, p-adjusted=0.515), on the level of rare and potentially damaging variants. Overall, the present study confirms the recent gene-based results for the implicated genes of the Xq28, distal locus in the latest meta-analysis of exome sequencing data in SCZ conducted by the SCHEMA Consortium. The small number of identified variants is partly explained by the strict filter criteria and limited sample size. These findings might indicate a possible contribution of ultra-rare variants locus and a high degree of conservation within this region. Moreover, this may point towards alternative pathomechanisms not investigated in the present analyses; such as the involvement of sex-dependent risk factors, a genuine overexpression of a disease-relevant gene or the disruption of topologically associated domains in the context of duplications at Xq28,distal. Although inconclusive, the present findings highlight the need for more research into the role of X-chromosomal genetic risk factors in SCZ.